A great deal of study involving various kinds of nanoparticles has recently and actively been conducted in biomedical fields such as cell staining, cell separation, in vivo drug delivery, gene delivery, diagnosis and treatment of disease or abnormality, molecular imaging, or the like.
In order to identify substantial significance of medical applications of such nanoparticles, satisfactory results should be achieved both in-vitro and in-vivo.
That is, nanoparticles with beneficial effects primarily proved through cell experiments, are then subjected to secondary animal testings to support that the tested nanoparticles so they may be applicable for medical use.
Magnetic resonance imaging (MRI) is a well known method for provision of anatomic, physiological and/or biochemical information of a human body through images by spin relaxation of hydrogen atoms in a magnetic field and is at present an excellent image diagnostic instrument that enables real time imagination of organs of an animal or human in a non-invasive way.
For precious and various utilizations of MRI in biological science or medical fields, a process of injecting a foreign substance into the body to increase contrast of an (MRI) image is used. In this regard, the foreign substance is often referred to as a contrast agent. Such a contrast agent may be a substance using super-paramagnetic or paramagnetic material that induces contrast of signals on a site to be observed through MRI, thus allowing the site to be clearly distinguished.
On MRI images, contrast between tissues is a phenomenon occurred due to difference in relaxation between tissues, wherein the relaxation refers to recovery of nuclear spin of water molecules in the tissues to an equilibrium state. The contrast agent influences such relaxation and thus may increase the difference in relaxation between tissues and induce variation in MRI signals, thus enabling clearly distinguishable contrast of the tissues. However, the contrast agent may cause differences in utility and precision, depending on the characteristics and functions of the contrast agent, subjects for injection of the contrast agent, or the like.
In addition, when contrast is improved using the contrast agent which helps to regulate image signals of specific organs and/or tissues to be higher or lower than adjacent organs and/or tissues, a more distinctive (sharp) image is created. A contrast agent increasing the level of image signals at a desired site of the body, from which MRI images are obtained, than that of the other site (adjacent to the desired site), may be referred to as a ‘positive’ contrast agent (‘T1 contrast agent’). On the other hand, a contrast agent decreasing the level of image signals at a desired site than that of the other side may be referred to as a ‘negative’ contrast agent (‘T2 contrast agent’). More particularly, the MRI contrast agent may be classified into a T1 contrast agent using high spin of a paramagnetic material and a T2 contrast agent using magnetic inhomogeneity around a paramagnetic or super-paramagnetic material. The ‘positive’ contrast agent relates to T1 relaxation, that is, longitudinal relaxation. Such longitudinal relaxation means that, after a magnetized component ‘Mz’ in Z-axis direction of the spin absorbs RF energy impact applied from X-axis, the magnetized component is aligned along Y-axis on an X-Y plane and emits energy to the outside, in turn returning to the original value (or state) of Mz. The foregoing action is expressed as ‘T1 relaxation.’ The time taken for returning Mz to 63% of an original value refers to “T1 relaxation time’ and, as the T1 relaxation time is decreased, MRI signals are greater, which in turn, decreases a period of time for acquiring images.
Likewise, the ‘negative’ contrast agent relates to T2 relaxation, that is, transversal relaxation. As described above, after the magnetized component ‘Mz’ in Z-axis direction of the spin absorbs RF energy impact applied from X-axis, the magnetized component is aligned along Y-axis on an X-Y plane and spontaneously decays and/or emits energy to adjacent spins, in turn returning to the original value of Mz. In this regard, another spin component ‘My’ equally widen on the X-Y plane is decayed by an exponential function and this is expressed as ‘T2 relaxation.’ A time taken until My is decayed to 37% of an original value refers to ‘T2 relaxation time’ and a My value measured through a receiving coil mounted on Y-axis by a function of time, wherein the My value is decreased over time, refers to a free induction decay (FID). Tissues with a short T2 relaxation time are shown as a dark region on the MRI.
In MRI contrast agents commercially available on the market, paramagnetic compounds are used as a ‘positive’ contrast agent while super-paramagnetic nanoparticles are used as a ‘negative’ contrast agent.
A current T2 contrast agent includes iron oxide nanoparticles such as SPIO (superparamagnetic iron oxide). In this case, T2 contrast is a negative contrast, that is, a negative contrast method wherein desired sites are darker than the surrounding part. Therefore, this method does not embody remarkable contrast effects and has a demerit of causing blooming effect to contrast a larger area than an actual size.
On the other hand, the T1 contrast agent has a merit of offering positive contrast to brightly display a desired site, and comprises a high spin material. Therefore, a gadolinium complex having 7 hole-spins in 4f orbital is usually employed. However, the gadolinium complex has very short in vivo and/or vascular retention time due to a relatively small molecular weight, causing difficulties in precisely diagnosing. Further, the above T1 contrast agent cannot be used to persons having weak kidneys because of a danger to derive nephrogenic systemic fibrosis and has recently received the warning by the U.S. Food and Drug Administration. Accordingly, there is a strong need for development of an improved T1 contrast agent that may solve such disadvantages of the gadolinium complex including, for example, short retention time, severe toxicity to patients with kidney diseases, or the like.
Among new trends in research on T1 contrast agents, an article regarding the use of manganese oxide nanoparticles having 5 hole-spins at 3d orbital has been disclosed (H. B. Na et al., Angew. Chem. Int. Ed. 2007, 46, 5397).
A manganese oxide nanoparticle has advantages such as a high T1 relaxation effect, which is a characteristic of manganese ions, and easy bonding to target molecules and easy intracellular injection which are characteristics of the nanoparticle. However, in the case where the manganese oxide nanoparticle is introduced into endosome, manganese ions escape from the nanoparticle due to internal acidic environments. Therefore, if such manganese ions remain in the body, these may cause a calcium channel disturbance problem (L. K. Limbach, et al., Environ. Sci. Technol. 2007, 41, 4158).
In order to overcome the above disadvantages, use of iron oxide as a T1 contrast agent, wherein the iron oxide has five hole-spins as well as higher biocompatibility than manganese, may be proposed.
General iron oxide (especially, magnetite or maghemite) nanoparticles are super-paramagnetic near room temperature. Due to such super-paramagnetic properties, that is, high magnetization, a T2 level is increased and susceptibility characteristics may occur, thus causing problems such as signal distortion. Consequently, it has been reported that magnetite is not suitable to be used as a T1 contrast agent (Y.-w. Jun, et al. J. Am. Chem. Soc. 2005, 127, 5732).
However, the foregoing problems may be overcome by controlling the size of iron oxide nanoparticles. More particularly, as the size of iron oxide particles is decreased, magnetic properties thereof may be reduced, which in turn deteriorates magnetic inhomogeneity. Accordingly, use of the iron oxide nanoparticles as a T1 contrast agent may be expected. For instance, U.S. Pat. No. 6,638,494 (inventor: Herbert Pilgrim) disclosed enhancement of T1 relaxivity (r1) by decreasing a particle size of super-paramagnetic iron oxide. According to the patent, iron oxide nanoparticles synthesized by co-precipitation, which have a particle size of 1 to 10 nm, and an average size (d50: median) of 2 to 4 nm, and hydrophilic surface, show that T1 relaxivity ranges from 2 to 50 L/mmol·sec and r2/r1 is 5 or less. However, although the particle average size (median) is small, a range of the particle size is considerably broad such as 1 to 10 nm, to thereby produce irregularity in particle size. If the size of an iron oxide particle is 4 nm or greater, the T2 effects may increase rapidly with the particle size. Therefore, even though the average size is small, improvement in T1 relaxivity is not so high when the particles have irregular size. Therefore, these nanoparticles are also not suitable to be used as a T1 contrast agent.
Among recent studies, use of iron oxide nanoparticles with a size of 4 to 6 nm as a T1 contrast agent has been reported (E. Taboada et al., Langmuir, 2007, 23, 4583; U. I. Tromsdorf et al., Nano Lett. 2009, 9, 4434). However, due to a relatively large particle size, T2 effects are still significant, thus the nanoparticles entail limitations in application thereof as a TI contrast agent.
Combidex® (AMAG Co.) which is currently under clinical trials in regard to use thereof as a T2 contrast agent for lymph nodes, had also been investigated for T1 contrast performance thereof. However, since an average size of iron oxide nanoparticles was relatively large in the range of 4 to 6 nm and a size of constitutional particles is irregular, it was known that T2 effect is predominant over T1 effect (Claire Corot et al., Advanced Drug Delivery Reviews 58 (2006) 1471).
Further, there is a method for preparation of iron oxide particles having a uniform size through thermal decomposition. However, strict requirements for the preparation of iron oxide nanoparticles having a size of 4 nm or less are needed, in turn not being preferable in commercial applications (Jongnam Park, et al., Nature Mater., 3(2004), 891).
Moreover, even if nanoparticles having a size of 4 nm or less may be prepared, raw materials are expensive and/or have severe toxic properties, thus having little significance in the aspect of commercial applications (Xiaowei Teng, J. Mater. Chem., 14(2004), 774).
Accordingly, synthesis and mass-production of iron oxide nanoparticles having a extremely small and uniform size of 4 nm or less, in a highly reproducible manner at low costs, as well as T1 contrast research using the same, have not yet been reported, which are still required.